To elucidate the process of fetal liver hematopoiesis, the relationships between stroma and hematopoietic cells involved in maturation were investigated. Cultured mouse fetal liver explants were established for morphological analysis of the interactions between fetal liver stroma and hematopoietic cells ex vivo. Fetal liver stroma comprised epithelial cells and macrophages, which occupied most of the culture surface. Macrophages proliferated extensively in primary culture, but almost disappeared after 3 passages. Close morphological and functional relationships were established between macrophages and hemopoietic cells, whereas epithelial cells did not interact with blood cells. Scanning electron microscopy revealed that macrophages were in close contact with erythroblasts and formed a three-dimensional network. In each erythroblastic island, 2-3 lymphocytes were also in contact with the macrophages; erythroblasts, lymphocytes and macrophages formed close, firm associations through their cytoplasmic membranes. This cell orientation was confirmed by transmission electron microscopy of fetal liver in vivo. In situ hybridization revealed that the macrophages expressed jagged-1, an important ligand of the Notch signaling system in hematopoiesis.